1. Field of the Invention
The present invention relates to dihydrodipicolinate synthase and dihydrodipicolinate reductase derived from thermophilic Bacillus bacteria and genes encoding the same.
2. Brief Description of the Related Art
In the production of L-lysine by fermentation, strains isolated from nature or artificial mutants thereof have been used to improve productivity. Many artificial mutant strains that produce L-lysine are known, and many of them are aminoethylcysteine (AEC) resistant strains and belong to the genus Brevibacterium, Corynebacterium, Bacillus, or Escherichia. Furthermore, various techniques have been disclosed for increasing amino acid production by such stains, for example, use of a transformant obtained by using recombinant DNA (U.S. Pat. No. 4,278,765).
Dihydrodipicolinate synthase (abbreviated as “DDPS” hereinafter) is an enzyme that synthesizes dihydrodipicolinate through dehydration condensation of aspartic acid semialdehyde and pyruvic acid, and this reaction serves as a starting point of the L-lysine biosynthesis system for the biosynthesis of aspartic acid-type amino acids. Furthermore, dihydrodipicolinate reductase (abbreviated as “DDPR” hereinafter) is known as one of the important enzymes of the Lysine biosynthesis system, and catalyzes the reaction in which the dihydrodipicolinate generated in the aforementioned reaction is reduced to generate piperidinedicarboxylic acid.
As for microorganisms belonging to the genus Escherichia or Corynebacterium, the gene (dapA) which codes for DDPS has been cloned, and the nucleotide sequence thereof has also been determined. As for the genus Escherichia, methods for producing L-lysine by enhancing DDPS have been disclosed in Japanese Patent Laid-open Publication (Kokai) No. 56-18596/1981, U.S. Pat. No. 4,346,170, and Applied Microbiology and Biotechnology, 15, pp. 227–331 (1982). Furthermore, a method for producing L lysine using an Escherichia bacterium into which DDPS derived from Corynebacterium bacteria has been introduced is known. DDPS derived from Corynebacterium is known to not suffer feedback inhibition by L-lysine, and has been disclosed in Korean Patent Publication No. 92-8382.
The gene coding for DDPR (dapB) has also been obtained from the genus Escherichia (Bouvier, J. et al., J. Biol. Chem, 259, 14829 (1984)) and the genus Corynebacterium (Journal of Bacteriology, 175(9), 2743–2749(1993)). Furthermore, a method for improving the production rate and production of L-lysine by enhancing the dapB gene derived from Corynebacterium bacterium together with the aspartokinase gene (WO96/40934) has been disclosed.
The current maintain of L-lysine production is fermentative production using a coryneform bacterium or an Escherichia bacterium. In this production, however, enzymes required for the fermentation may be inactivated or the production bacteria may be killed due to a temperature increase in the medium during the fermentation, and thus it is necessary to cool the medium during the fermentation.
Enzymes and proteins produced by thermophilic bacteria are generally stable at elevated temperatures, and are also stable against pH variation or organic solvents. Therefore, applications thereof as diagnostic regents, industrial catalysts, and so forth have been highly developed. If it is possible to produce L-lysine by fermentation at elevated temperatures by utilizing stable and durable enzymes derived from thermophilic bacteria, cooling of the medium becomes unnecessary and thereby reduces the cost. Moreover, if fermentation at elevated temperatures is realized it is expected that the reaction rate may also be improved